RU2408361C2 - Local nail compositions - Google Patents

Abstract

FIELD: medicine, pharmaceutics. ^ SUBSTANCE: group of inventions refers to medicine, namely to dermatology, and can be used in pathological state of nails. A method of treating wherein a first composition containing a reducing agent of potential more than +20 mV and a second composition containing an oxidising agent of potential less than -50 mV are used. The compositions are applied to coat a nail separately consistently with the first composition applied in the beginning, and then the second composition. Either the first, or second, or third compositions which is a separate option for the compositions with the restoring and oxidising agents, containing a medicinal agent. Also, there is offered a kit comprising the first and second compositions which are arranged separately, and the medicinal agent which is included either in the first composition, or in the second composition, or optionally in the third composition. ^ EFFECT: inventions improves clinical effectiveness in the pathological state of nails due to intensified penetration of the medicinal agent through a nail plate so that the effective concentration of active particles reaches deeper layers of the nail plate, and also the same nail bed. ^ 45 cl, 9 tbl, 11 dwg, 10 ex

Description

The present invention relates to topical compositions for nails, compositions comprising a drug and a penetration enhancer.

Onychomycosis is a general term for fungal infections of the nail plate or nail bed, and is the cause of 50% of nail diseases. Both fingernails and toenails can be affected. It is estimated that approximately 5% of the population is currently affected in Europe and the disease is becoming more common. This increase in prevalence is mainly associated with an aging population, since onychomycosis is much more common in older people. Other predisposing factors include bad shoes and an increase in the frequency of use of public areas for recreation.

The pathogens most often responsible for the development of onychomycosis are dermatophytes, which are believed to be the cause of more than 90% of all cases of the disease. Trichophyton rubrum (56% toenails - 36% toenails) and Trichophyton mentagrophytes (19% toenails - 11% toenails) are especially common. Yeast infections are less common, but are usually associated with Candida albicans (10% to 30% toenails).

The second, relatively common nail disease is psoriasis. Psoriasis is best known as an inflammatory skin disease, but most patients who suffer from psoriasis of the skin also suffer from nail psoriasis. Patients with only nail psoriasis are rare. Psoriasis is most common in Europe and North America, where it affects about 3% of the population.

Despite the fact that nail diseases are rarely life-threatening, they can be very painful and disfiguring for the patient. The main symptoms include a discoloration of the nails, often to a yellow / green or darker color, and the accumulation of organic residue under the nail, causing an unpleasant odor. In addition, the nail may thicken and exfoliate. Such aesthetic manifestations themselves can have a serious impact on the quality of life of the patient. In addition to these symptoms, the general condition can be very painful. In particular, thick toenails can cause discomfort when wearing shoes and may even cause inconvenience to some patients when standing and walking.

The effective treatment of onychomycosis, and other diseases of the nails, is seriously hindered by the fact that the infected area is effectively protected by the nail plate, which consists mainly of keratins, a fibrous group of proteins. Keratin fibers are held together by cysteine-rich globular proteins whose disulfide bonds act like glue and are responsible for most of the integrity of the nail plate. Any effective method of treatment should be able to overcome the obstacle represented by a rigid and solid nail plate, and deliver active particles to the nail bed.

Known methods of treatment are divided into three main categories. The first involves removing all or part of the affected nail to affect the infected area. Removal may be surgical or chemical. Chemical removal may include applying urea to the nail plate within a tight dressing for a short period. The action of urea is based on the breaking of bonds between proteins in the nail. The nail becomes soft and separates from the nail bed. However, the traumatic and painful nature of such treatment means that it is unpopular, and is used only as a last resort.

The second main method of treatment is the oral administration of the appropriate drug. There are currently four major oral therapeutic agents available for the treatment of onychomycosis. These include griseofulvin (Grisovin®, GSK), ketoconazole (Nizoral®, Janssen-Cilag), itraconazole (Sporanox®, Janssen-Cilag) and terbinafine (Lamisil®, Novartis). For psoriasis, oral agents such as methotrexate, etretinate, and cyclosporine may be effective.

Griseofulvin has been available since the 1950s. Due to its fungistatic activity against dermatophytes, long treatment periods are required (9-12 months for toenail infections). Cure rates are low and relapse rates are high. Ketoconazole was the first imidazole-based drug to be included in the treatment of onychomycosis in the 1980s. However, due to its hepatotoxicity, its use is now limited to nail infections that do not respond to other therapies. More recent antifungal agents, itraconazole and terbinafine, are more effective in the oral treatment of onychomycosis, with higher mycological indicators of treatment effectiveness and shorter treatment periods than previously observed. The third method involves topical application of the composition to the nail plate. Topical commercially available onychomycosis treatments include amorolfine (Loceryl®, Galderma) and cyclopirox (Penlac®, Dermik). The terms "local" and "topical" as used herein mean application to a surface such as skin or nail, in contrast to systemic use, which is usually carried out by ingestion or injection.

Topical treatments for nail diseases are mostly preferred because they do not have the risks of systemic drugs such as hepatotoxicity and are less painful and disfiguring than treatments involving total or partial removal of the nail. However, in order to have an effective effect, the active particles must be able to penetrate the nail plate in sufficient quantities so that the effective concentration of the active particles can reach the deeper layers of the nail plate, as well as the nail bed itself. It is likely due to insufficient drug penetration that currently available topical treatments are relatively ineffective, and are associated with long duration of treatment and low rates of treatment effectiveness.

Compared to a thin stratum corneum, the nail plate is much thicker. This means that there is a much longer path of entry for drug delivery to the nail bed. In addition, the nail does not act as a lipoid barrier, but rather as a concentrated hydrogel. The disulfide bonds of cysteine-rich proteins are largely responsible for the integrity and structure of the nail and its barrier properties. Thus, the development of effective local treatments for nail diseases is a much bigger task than the development of local skin care products.

There are several factors that affect the rate of penetration of drugs into and through the nail plate. These factors include the size of the penetrating particles, the hydrophilicity / lipophilicity of the penetrating particles, and the nature of the carrier. Additionally, penetration can be enhanced by effectively reducing the barrier that drugs must penetrate to reach the site of infection in the nail bed. The barrier can be reduced physically or chemically.

Physical reduction of the barrier involves the partial or complete removal of the nail, or filing the upper layer of the nail plate. Such procedures are undesirable, as they are both painful and disfiguring. A more acceptable approach involves the use of chemical enhancers that, when applied to the nail, interact and modify the structure of the nail, so as to reduce the barrier to the penetration of the drug and increase the diffusion rate of the active particles in and through the nail plate. Urea is usually used as a penetration enhancer for nails, due to the fact that it has the ability to chemically remove nail plates. Other approaches focus on disulfide bonds in the nail, and the disintegrants of these bonds, including acetylcysteine and mercaptoethanol.

Currently, there are no local topical treatments available that show satisfactory results in treating conditions affecting the lower layers of the nail plate and nail bed.

US 6,664,292 discloses topical compositions for treating pathological conditions of nails. The compositions are composed of lower alcohol and lower carboxylic acid.

US Pat. No. 5,753,256 discloses a patch for treating nail mycosis. The patch contains an active compound and a penetration enhancer. Penetration enhancers such as sulfoxides, lactic acid, salicylic acid, propylene glycol, dimethylformamide, dimethylacetamide and sodium dodecyl sulfate are disclosed.

US Patent No. 2003/0235541 discloses an aqueous, basic composition for topical treatment of onychomycosis.

US Patent Application No. 2001/0049386 discloses a method for treating onychomycosis in which a softening composition comprising urea and an antifungal composition is administered to the infected area around the nail, either in one or in separate compositions, simultaneously or simultaneously.

WO 99/40955 discloses a patch with a pressure sensitive adhesive matrix for the treatment of onychomycosis. The patch does not necessarily contain skin penetration enhancers.

GB 2278056 A discloses thioglycolic acid higher esters and amides as penetration enhancers for use on the skin, and compositions suitable for the treatment of onychomycosis are disclosed. However, these compositions can only be applied periungal. In addition, this treatment still requires the systemic presence of the drug, as well as compositions intended for transdermal administration, so that despite the fact that the therapeutic agent can be applied locally, the drug will still enter the bloodstream.

In patent WO 99/49895 it is disclosed that thioglycolic acid is able to restore keratin in the nail and, therefore, can be used to improve the diffusion of drugs through the nail to the nail bed.

DE 1000 567 discloses the use of thioglycolic acid in combination with sodium iodide to restore keratin in a nail, while EP 0712633A1 simply discloses the use of thioglycolic acid as a penetration enhancer for the skin.

US 2003/0007939 discloses a pharmaceutical composition of hydrogen peroxide and at least one other dermatological agent to enhance penetration into the skin, scalp, hair and nails. The use of a reducing agent is not described.

EP 0425507 discloses a composition for treating an epithelium, including cutaneous, in a condition of pathology or damage, which comprises an activated protein, an oxidizing agent, including hydrogen peroxide, and a reducing agent, including thioglycolic acid.

Thus, there is still a need for effective topical compositions for treating pathological conditions of nails.

Currently, to our surprise, we have found that a reducing agent such as thioglycolic acid (TA) and an oxidizing agent such as hydrogen peroxide can be applied to the nail one by one to enhance the penetration of the drug. Under ordinary circumstances, a combination of these two agents generally results in an extreme chemical reaction. However, combining these agents on the nail was safe.

Thus, in a first aspect, the present invention provides the use of a composition, preferably a liquid, of each reducing agent and an oxidizing agent in the manufacture of a medicament for treating pathological conditions of medically treatable nails, said compositions are used separately and when applied sequentially to a patient’s nail in the order in which a reducing agent is applied followed by an oxidizing agent, and where the drug is used in one or the other composition, or optionally in a third composition, in which the drug is complementary to the reducing or oxidizing agent. Any or a specific reducing agent and / or any or specific oxidizing agent may be selected from known drugs, such as antifungal and antipsoric drugs, which have suitable reducing or oxidizing properties, but it is generally preferred that an additional drug is used as shown below .

Preferably, the reducing and oxidizing agents are powerful enough so that a simple combination of these two drugs leads to an extreme reaction, such as a mostly exothermic, or even explosive, reaction. Now, surprisingly, it has been found that when applied to the nail in a sequential manner, when a reducing agent is applied, followed by the application of an oxidizing agent, such combinations can be safely used without immediate visible signs of an extreme reaction. It is clear that some reducing and oxidizing agents will be too powerful to be applied safely to the nail, or lead to undesirable reactions, such as discoloration, which may be undesirable for some patients. However, provided that each agent individually is acceptable for application to the nail, combinations are also acceptable, even if a combination of these two agents, in the absence of a nail, would lead to an extreme reaction. Agents such as thioglycolic acid and hydrogen peroxide are an example of one such combination. Other agents / combinations can be easily selected by specialists in the field of technology, using the above criteria.

Suitable concentrations for each agent are independently selected, and they generally comprise from 1 to 50% v / w, with a preferred range of 5 to 30%. More preferably, the concentration of each is selected separately from a range of from 10 to 25%, and especially from 5 to 15%.

In a preferred embodiment, the desired strength of the reducing and oxidizing agents can be selected based on their reducing potentials. As is known in the art, the reduction potential of specific particles indicates the ability of these particles to attach electrons, thus providing data on their ability to recover.

The reduction potential of the oxidizing and reducing agents according to the present invention can be determined in accordance with any procedure known to those skilled in the art. A preferred method is to determine the reduction potential of a solution of a particular agent in water or in a mixture of water and ethanol, relative to a silver / silver hydrochloric acid electrode, in r.t.p. and calibrated by standard redox potentials.

According to a preferred embodiment of the invention, the reduction potential of the oxidizing agent, as determined according to the aforementioned protocol, is preferably less than -50 mV, more preferably less than -100 mV, and most preferably less than -200 mV. The reduction potential of the reducing agent is preferably more than +20 mV, more preferably more than +50 mV, and most preferably more than +75 mV.

As shown in the accompanying examples, the reduction potential of solutions containing each agent can be adjusted by changing the concentration of the oxidizing or reducing agent present in the solution, or by adjusting the pH of the solution, if necessary.

Thus, in a preferred embodiment, the reducing agent is made in the form of an alkaline composition with a pH between 7 and 14. More preferably, the pH is from 8 to 13, with a particularly preferred pH from 9 to 12, since this mainly maximizes the reduction potential of the reducing compound .

On the other hand, while the pH of the oxidizing agent is generally lower, it is preferable that the pH is from 1 to 7, with a more preferred pH between 2 and 5, since this leads to a maximum increase in the oxidation potential (-ve reduction potential ) In any case, it is preferable that no composition is so acidic / corrosive that careless spraying on the skin does not lead to pharmaceutically and / or cosmetically undesirable damage.

However, it is clear that the pH of each composition can be changed in order to achieve the desired effect, for example, to reduce the reduction potential or the reaction rate, so that the above preferences are applied mainly when there are no other significant pH considerations.

In general, the compositions of reducing and oxidizing compounds are selected so that the reducing potentials are within the ranges indicated above, and mainly to maximize the + ve or -ve potential, if necessary. While it is generally preferred that the reduction potential is optimized by concentration, it is clear that compositions having low concentrations can be provided in which evaporation of the solvent and / or co-solvent will result in higher in situ concentrations. Preferred concentrations of agents are readily ascertained in accordance with the methods given in the accompanying examples. For example, a preferred concentration range of thioglycolic acid is at least 0.1% to 20% and higher; urea-H ₂ O ₂ at least 5% to 40% and higher, H ₂ O ₂ at least 20% to 100%, and DTT at least 0.05% to 20% and higher. Compositions containing ethanol or another volatile solvent or co-solvent can be made with lower concentrations of a reducing or oxidizing agent, since the concentration of the agent will increase when applied to the nail, when the composition is also applied to it.

It is particularly surprising that the use of these two components appears to be synergistic, in the sense that there is no reason to suspect that the increased permeability caused by any component is different enough to be complemented by the other. Indeed, this method of exposure is not expected in the technical field, since the use of an oxidizing agent in the first place mainly does not lead to any significant improvement in the permeability of the nail more than that achieved with a more efficient use of the two components. However, the use of a reducing agent in the first place, apparently, leads to such synergy that often a greater effect is achieved than with the combined action of both agents. This effect can be, but not always, reflected to some extent in increasing the absorption of fluid by the nail (underneath).

Preferred reducing agents include ammonium thioglycolate, calcium thioglycolate, sodium thioglycolate, thioglycolic acid (TA) and dithiothreitol (DTT), ascorbic acid, hydroquinone, mercaptoethanol, glutathione, L-cysteine, taurine, aminomethanesulfonic acid, cysteine sulfonate acid, cysteine sulfate acid, cysteine sulfate acid, cysteine sulfate acid, cysteine sulfate acid acid, ethanesulfonic acid, homotaurin, hypothaurine, isethionic acid, mercaptoethanesulfonic acid, N-methyltaurine (MTAU), as well as their simple derivatives.

By "simple derivative" is meant a salt, ether or amide of a compound. Simple derivatives of thioglycolic acid are particularly preferred. Any other alkyl component is preferably lower alkyl having only 1 to 6 carbon atoms, but more preferably having 1 to 4 carbon atoms, and most preferably 1, 2 or 3 carbon atoms.

The reducing agent is preferably thioglycolic acid or a derivative thereof and, in fact, a reducing agent as such is usually referred to here, although it should be understood that any such reference will also include other reducing agents, unless it is clear or obvious otherwise.

The oxidizing agent may be any compound that is suitable, including urea, hydrogen peroxide, potassium persulfate, thiouracil, p-coumaric acid, glycolic acid, oxalic acid, cineole, peroxidone, chlorine dioxide, ammonium dichromate, ammonium nitrate, ammonium perchlorate, permanganate ammonium, barium bromate, barium chlorate, barium peroxide, cadmium chlorate, calcium chlorate, calcium chromate, calcium perchlorate, chromium nitrate, cobalt nitrate, silver oxide, periodic acid and pyridine dichromate. Hydrogen peroxide is preferred, and the product of the compound of hydrogen peroxide and urea is more preferred.

The compositions can be administered immediately one after the other, but it is preferable to apply the compositions in the order of application of the reducing composition followed by the application of the oxidizing composition and allowing the first composition to react with the nail for a short time before applying the oxidizing composition. A suitable time is from 10 seconds to 10 minutes, more preferably from 1 minute to 5 minutes. Although such short periods of time are acceptable, it has been found that the applications can be separated by significant periods of time, and the preferred time interval between the application of the reducing and oxidizing agents is from 15 to 30 hours, and more preferably from 20 to 26 hours, with similar periods between subsequent applications. When such longer periods of time are maintained, it is preferred that the drug is present in one or both of the compositions, or is administered in a separate composition, but together with one or both of the drugs.

The drug may be present in one or both of the compositions, or may be prepared separately for administration before, during, or after the application of oxidizing and reducing agents. In one embodiment, a reducing agent is applied to the nail, followed by application of the drug and then the oxidizing agent, so that the drug is in place when the interaction between the oxidizing and reducing agents leads to improved penetration. It is generally preferred that the drug is in a separate composition, especially when prolonged exposure to one or both of the oxidizing and reducing agents is undesirable.

The compositions of the present invention are intended for application to nails, and can be made in any form suitable for application to nails. The compositions are preferably aqueous, optionally with a co-solvent such as ethanol or acetone. Although a co-solvent may not be required for oxidizing or reducing agents, it may be necessary to dissolve the drug.

In general, the nail compositions of the present invention can be provided in the form of creams, ointments, gels, solutions, lotions, foams, mousses, sprays, pastes or varnishes when they are intended for direct application to nails. However, compositions of the invention may also be provided, for example, in the form of solutions or powders or pre-prepared mixtures. For example, the solution can be applied to a dressing, such as a patch, and then connected to the nail to more accurately get to the desired area. Alternatively, the dressing can be applied to the nail, and then the solution is applied to the dressing. In this case, for example, it is possible to limit the dressing only to the surface of the nail, or the dressing can be made in such a way that the contact between the solution and the surfaces on which there is no nail, or the area being treated is suppressed or prevented, or by means of a design dressing, or through the introduction of insulating agents. Suitable insulating agents may include some adhesives or resins, or other suitable agents. In the case of the use of such dressings, it is preferable that for each reducing and oxidizing agents, separate dressings are used to avoid the possible development of an explosive reaction.

When a dressing is provided, it may also be desirable to apply a further occlusive dressing to the dressing if a solution or other drug composition has been applied to the absorption dressing to prevent evaporation of the carrier when this is not desired.

For example, a patch similar in construction to a transdermal patch, but preferably a reservoir patch, can be used to provide one agent after applying another in the form of paint or varnish. Similarly, it is possible, for example, to apply the powder by sprinkling on a varnished or painted surface.

Other powders and pre-prepared mixtures can be further manufactured, if desired, for example, in the form of solutions or compositions for applying to nails.

You must understand that references to "nails" here include links to any suitable nail surface. In humans, this will actually only include fingernails and toenails, but any nail surface is contemplated. Suitable carriers for agents mainly include water and lower alcohols, including monohydric and polyhydric alcohols. Other solvents may also be used, such as acetone and, in general, it will be apparent to those skilled in the art which solvents are suitable for application to nails.

In addition to the carrier, other bases can be used, for example, such as bulking agents. Although water may be used to form the volume of the composition, it has been found that propylene glycol is generally associated with a higher degree of penetration into the nails, and is generally a more preferred bulking agent.

The compositions may be simple aqueous compositions, or may contain other components, such as, for example, thickening agents, stability enhancers, pH modifying agents, odor inhibitors, and colorants. The use of odor inhibitors may be particularly preferred when the drug is intended to treat a pathological condition of the nail accompanied by a fetid odor, which is often found in such conditions, or when it is desirable to mask the smell of the drug directly. Similarly, dyes may be included when the pathological condition being treated leads to an undesired discoloration of the nail. In one embodiment, the compositions are formulated as varnishes that dry in situ.

In another embodiment, reducing and / or oxidizing agents, and especially thioglycolic acid, can be conjugated to the drug used.

By the term “drug” is meant any active ingredient of a composition of the invention that is capable of exerting a therapeutic effect when applied to a nail. The therapeutic effect may be noticeable only in relation to the penetration enhancer of the invention.

Suitable drugs for use in the compositions of the invention may be suitable for any pathological condition associated with a patient’s nails, but will often fall into the category of either mycosis or a condition associated with psoriasis. The drug may be presented in any suitable form, and may be a solid, liquid, or gas present in the composition for application to the nail. Suitable gases include, for example, NO.

Suitable drugs for use in the compositions of the present invention include antifungal drugs: amorolfine, miconazole, ketoconazole, itraconazole, fluconazole, econazole, cyclopirox, oxyconazole, clotrimazole, terbinafine, naftifin, amphotericin, grisiefin-isolofin, isofin-isofinolin isofin-isolofin, isofin-isolofin-isolofin-isolofin-isolofin-isolofin-i-isofin-olivine-iolofin-isolofin-isofin-azolefin-iolinfin-azolefin-iolofin-azolefin-iolin-azolefin-olivine-isolofin-isolofin-isofin-olifin-olin-isofin-iolofin-iolofin-azolefin-iolofin-azolefin-iolofin-i-solumin and ethers. Terbinafine is particularly preferred.

Suitable drugs for use in relation to psoriasis include corticosteroids, 5-fluorouracil, methotrexate, etretinate, cyclosporin, tacrolimus and their derivatives.

The amount of drug used is not limited to the invention, and it is only required that the drug can be used in an effective amount for the treatment or prevention of a pathological condition of the nail. The amount of the drug may depend on the age, gender and / or weight of the patient, but will mainly be provided in the form of a finished composition for application to the nail, and the concentration of the drug will mainly depend on the pathological condition being treated as the main determining factor.

The amount of the drug may be sufficient to eliminate the pathological condition after one use, but basically there is a continuation of the treatment in the form of multiple applications, so that the amount of the drug can be adapted for gradual treatment according to the intended number of cycles.

In cyclic treatment, the treatment of an infected nail is carried out by a cyclically reducing agent and an oxidizing agent. A preferred strategy is to treat with a reducing agent on day 1, then an oxidizing agent on day 2, with a drug that is preferably administered with or shortly after the oxidizing agent, and then repeating the process on day 3, starting with applying the reducing agent again . Optionally, a gap may be provided between treatments so that re-treatment begins again, for example, on day 4, day 5, day 6, or day 7. The period may be longer, if preferred, but preferably not more than a month, and preferably not more than 2 weeks.

It is clear that the cycle may include two or more applications of any of the compositions. For example, a reducing agent can be used every day, for example, for two or three consecutive days, or several times in one day, with an oxidizing agent and a drug used on the day after the last treatment with the reducing agent, or one day after the first treatment, which may be shortly after the last treatment with a reducing agent. The oxidizing agent may be used on consecutive days, or several times over the course of one day, and it is generally preferred that the drug is administered simultaneously or shortly after each treatment with the oxidizing agent. Suitable concentrations of the drug in the compositions of the invention will depend on factors such as the pathological condition being treated and the drug to be used, and, in any case, will be readily apparent to those skilled in the art. For information, suitable concentrations may be in the range from 0.1% to 50% v / v, more preferably from 1% to 20% v / v, especially from 1% to 10% v / v, despite the fact that concentrations both above and below these ranges are provided in accordance with the present invention.

The compositions of the present invention may further include other drugs and / or penetration enhancers. Suitable examples of various drugs are presented above. Other penetration enhancers include lactic acid, DMSO, salicylic acid and oleic acid, of which lactic acid, salicylic acid and oleic acid are individually preferred.

The concentration of the enhancer can be any that is effective to allow greater penetration into the nail plate than a similar composition containing no enhancer. In general, the amount of amplifier varies from about 0.1% v / v. up to about 25% v / v, with amounts from about 1% to 20% and, more preferably, from 3% to 15% v / v, often providing good results.

The present invention also provides a method for treating a nail infection in a patient in need, comprising applying a reducing agent composition to said nail, followed by applying an oxidizing agent composition thereto, said pathological condition of the nail is treatable with a medicine, said medicine is in one of said compositions or in a third composition.

You must understand that the above method preferably includes any properties that are defined above with respect to application, and / or which are defined in any of paragraphs 2 to 43 of the attached claims.

The present invention further provides a kit comprising compositions as defined above for treating a nail infection, and preferably as defined in any of the claims 1-43 of the claims.

The present invention will now be further illustrated by the following non-limiting examples.

Examples

Examples 1-4

Materials and methods:

In Examples 1-4, two models were developed to determine the permeability of the nail. The first model corresponded to a mode similar to that described in WO 99/49895, in which the nail was treated with a penetration enhancer dissolved in a solvent, and the% increase in the weight of the nail was determined. It is believed that penetration enhancers are keratolytic and, thus, break the bonds between the sulfur atoms in the nail, thus causing it to soften and absorb more fluid.

The second model actually evaluates the permeability of the nail. In this new model, a radioactive compound, ¹⁴ C-mannitol, was used to track the progress of the agent through the nail after application of penetration enhancers. To assess the diffusion of mannitol, a new diffuser was used.

In examples 1-4, instruments, materials and methods are used, which are described in detail below (table 1).

Materials:

Table 1 Suppliers, purity and other details
about the materials used in the study Product Provider Address Purity Ammonium Thioglycolate Fluka Sigma-Aldrich Company Ltd, Dorset, UK ≈60% in water Calcium thioglycolate Fluka Sigma-Aldrich Company Ltd, Dorset, UK ≥90% EDTA Sigma-alldrich Sigma-Aldrich Company Ltd, Dorset, UK 99% Ethanol Bdh BDH Laboratory Supplies, Poole, Dorset, UK 99.7-100% (Anala®Grade) 1,4-dithio-DL-threitol Fluka Sigma-Aldrich Company Ltd, Dorset, UK Molecular Biology Purity ≥99% Glycolic acid Sigma-alldrich Sigma-Aldrich Company Ltd, Dorset, UK ReagentPlus ™
99% Hydrogen peroxide solution Sigma-alldrich Sigma-Aldrich Company Ltd, Dorset, UK 35 wt.% In water Hydrate L-cysteine hydrochloride Sigma-alldrich Sigma-Aldrich Company Ltd, Dorset, UK 99% Sodium thioglycolate Sigma-alldrich Sigma-Aldrich Company Ltd, Dorset, UK 96.5-103.5% by titration with iodine Thioglycolic acid Sigma-alldrich Sigma-Aldrich Company Ltd, Dorset, UK ≥98% Hydroperite Addition Compound Sigma-alldrich Sigma-Aldrich Company Ltd, Dorset, UK 98% Water Millipore Millipore (U.K.) Limited, Gloucestershire Milli-Q Gradient System (18.2MΩ-cm Resistance) ³ H water MB11 material 37MBq / ml Sigma-alldrich Sigma-Aldrich Company Ltd, Dorset, UK ³ H water MB11 material 37 MBq / ml 14C mannitol CFA 238 material 9.25 MBq in 1.25 ml GEC Amersham Pharmacia GES Amersham Pharmacia. St. Giles Bucks 14C mannitol CFA 238 material 9.25 MBq in 1.25 ml

Penetration Amplifiers:

Since each of the penetration enhancers showed a different solubility, either water or a mixture of ethanol and water was used as solvents, in accordance with Table 2 below.

table 2 Solvents used for
penetration enhancers Agent Solvent Ammonium Thioglycolate 20% ethanol Calcium thioglycolate water Sodium thioglycolate 20% ethanol Thioglycolic Acid (TA) 20% ethanol Glycolic acid 20% ethanol Dithiothreitol (DTT) 20% ethanol Cysteine 20% ethanol Urea-H ₂ O ₂ water

Methods

Nail swelling study:

Washing and preparing pieces of nail:

Nail pieces of approximately 2 mm in length are obtained from healthy volunteers (with their consent) using nail scissors. They were washed using 70% ethanol (v / v), using vigorous stirring in a 28 ml glass tube, using WhirliMixer ™ (Fison's, UK) at a maximum speed of 1 minute. Then the pieces are rinsed in water with vigorous thorough mixing for 1 min. This procedure is repeated three times. Then the nails are placed in an open Petri dish and left to dry in an oven with a controlled temperature of 30 ± 2 ° C for 24 hours. After drying in the oven, the nails are weighed and placed in individual wells in a 24-well microbiological plate (Costar®, UK). In all experiments, 10 sets of nail pieces were used.

Application of penetration enhancers to nail pieces:

The washed nail pieces in the wells of the microbiological plate are placed in 1 ml of a penetration enhancer solution for approximately 20 hours. Excess solution is removed from all pieces of the nail by gently wiping them dry with cloth. The nail pieces are weighed and the results of the weight measurement are recorded. If a second penetration enhancer is used, then the same procedure is repeated with the second compound and the second weight measurement result is recorded. To evaluate the effect of the solvent individually, a number of control nails are determined using an identical methodology, but simply by immersing them in a solvent without adding a penetration enhancer.

Nail penetration studies of mannitol:

Getting penetration enhancers

Separate penetration enhancers are prepared in a suitable solvent and labeled with ¹⁴ C mannitol (10 μl from the original ¹⁴ C mannitol). If more than one penetration enhancer is applied to the nails, only the last penetration enhancer is labeled with ¹⁴ C mannitol prior to application to the nail. ¹⁴ C mannitol without penetration enhancer is used as a control.

Preparation of a diffuser for nails:

Pre-calibrated nail diffusers fill a person’s fingernail with all thickness. The receiver fluid, which consists of a mixture of 20% ethanol / water, is placed in the receiver well to the engraved mark on the side arm. A magnetic follower is inserted and then the cells are placed on a magnetic stirrer in a water bath and the temperature is maintained at 32 ° C. Cells are checked for leakage and the formation of air bubbles. After a 30 min equilibration period, a 1 ml sample is taken from the side lever for sampling and placed in a scintillation tube, followed by 4 ml of a scintillation cocktail, and tested on a scintillation counter using a dual mode setting ( ³ H / ¹⁴ C) . This is done to ensure that no cells have any residual radioactivity, and these readings were also taken as a baseline for each cell. Then, the cells are poured to the engraved mark with fresh receiver liquid, previously balanced at 32 ° C.

Normalization of the nail using ³ H of water:

After obtaining the background data (described in the paragraph above), 50 μl of ³ H water (approximately 18000 dpm) is placed on top of the nail in the assembled nail diffuser using a pre-calibrated Gilson pipette. Then the cells are immediately covered with parafilm. After 20 hours, a 1 ml receiver fluid sample is removed from the side arm using a 1 ml syringe and placed in a scintillation tube followed by 4 ml scintillation cocktail and tested on a scintillation counter using a dual mode setting ( ³ H / ¹⁴ C) . This value is used to normalize each nail and, if it is believed that the radioactivity is too high, then the cells are rejected due to leakage. To determine the leakage from any cell, the Dixon Q-test is used. Then ³ H water is removed using a paper towel, and the cell is poured to the engraved mark with fresh receiver liquid, previously balanced at 32 ° C.

When testing a single penetration enhancer, aliquots of 50 μl of each penetration enhancer (pre-labeled with ¹⁴ C mannitol as described above) are pipetted using a pre-calibrated Gilson pipette onto the surface of a person’s fingernail of all thickness. Then each cell is covered with paraplenka. Sampling is performed at time points of approximately 20 hours, 44 hours, 68 hours, 92 hours and 116 hours. After each point in time, a 1 ml sample is taken from the side arm and placed in a scintillation tube, followed by 4 ml of scintillation cocktail and tested on a scintillation counter using a dual mode setting ( ³ H / ¹⁴ C). Then the cell is poured to the engraved mark with fresh receiver liquid, previously balanced at 32 ° C. Each penetration enhancer (and control) is tested a total of n = 5-6 times using the procedure presented above.

When using more than one penetration enhancer, 50 μl of a penetration enhancer without radioactive mannitol is pipetted using a pre-calibrated Gilson pipette onto the entire surface of a person’s nail, then each cell is closed with paraplenka. After 20 hours, the pre-penetration enhancer was removed with paper towels and the nail surface was washed with deionized water (3 × 1 ml). A second penetration enhancer, previously labeled with ¹⁴ C mannitol, is then applied, and then the protocol described above is applied to the separate penetration enhancer.

Example 1 :

The use of separate penetration enhancers for nails

Nail swelling studies:

In FIG. Figure 1 shows the change in the weight of the nail after 20 hours when water (control), cysteine, ammonium thioglycolate (AmTA), thioglycolic acid (TA), glycolic acid (GA), hydrogen peroxide (H ₂ O ₂ ), hydroperite ( Urea - H ₂ O ₂ ) and 1,4-dithio-DL-threitol (DTT) (value ± standard deviation, n = 10). Values are expressed as a percentage of the initial weight of the nail. Despite the fact that both water and 20% ethanol in water were used as control solutions, no significant difference was observed between them (p> 0.05, ANOVA). Therefore, figure 1 shows only water control.

Thioglycolic acid (TA) 5 glycolic acid (GA), hydrogen peroxide (H ₂ O ₂ ), hydroperite (urea-H ₂ O ₂ ), dithiothreitol (DTT) and ammonium thioglycolate (AmTA) all significantly increased (p <0.05 , ANOV A) the weight of the pieces of the nail after 20 hours compared with the control. The increase in nail weight caused by six of the seven penetration enhancers tested means that the nails absorb more fluid applied. A physical change in the appearance of the treated nails was also noted. The nails treated with TA, GA, H ₂ O ₂ and AmTA were softer than the control samples and also had a lighter color.

Cysteine did not significantly increase (p> 0.05, ANOVA) the weight of the nail. Thus, the penetration of fluid into the nails treated with cysteine was not enhanced.

Studies of penetration into the nails of mannitol:

In FIG. 2 shows the penetration of ¹⁴ C mannitol through a human nail of entire thickness using various individual penetration enhancers (n = 6 ± standard deviation, DTT n = 5 ± standard deviation). Saturated TA (50%) was found to be the best mannitol permeation enhancer in these experiments. However, when TA was restored to a concentration of 5%, its penetration enhancing effects decreased lower than that of DTT. H ₂ O ₂ does not differ significantly from 5% TA (p <0.05, ANOVA) with respect to diffusion of mannitol after 96 hours.

Example 2: Application of binary penetration enhancers NaU

Nail swelling studies:

The maximum increase in nail weight when applying a separate penetration enhancer was 71% for 5% TA (shown in Fig. 1). However, the addition of two penetration enhancers, one after the other, led to an increase in the weight of the nail up to 150%, as shown in FIG. 3. In FIG. Figure 3 shows the increase in the weight of nails in percent when treated with 2 penetration enhancers at 2 separate stages (> indicates the next step) in both application modes (value ± standard deviation, n = 10).

The addition of TA and then H ₂ O ₂ or urea-H ₂ O ₂ significantly increased the weight of the nail (p <0.05, ANOVA) compared to TA alone. However, when H ₂ O ₂ or urea-H ₂ O _{2 was} applied before TA, this did not significantly increase the weight of the nail (p> 0.05, ANOVA) compared to TA alone, a similar tendency was observed in each of the experiments with double penetration enhancers presented in detail in FIG. 3. The addition of a reducing agent (for example, TA, GA, DTT) before the oxidizing agent (for example, H ₂ O ₂ or urea-H ₂ O ₂ ), significantly increased the increase in the weight of the nails compared to applying the agents in the reverse order, that is, oxidizing and then reducing agents. The application of TA and then urea-H ₂ O ₂ caused the greatest weight gain, while the least was observed when applying urea-H ₂ O ₂ and then DTT.

Example 3 : Effect of the concentration of penetration enhancer

Nail swelling studies:

In FIG. Figure 4 shows the increase in the weight of nails in percent after a period of 20 hours at various concentrations of TA (value ± standard deviation, n = 10). In the case of nails treated with TA, an increase in the concentration of the penetration enhancer resulted in increased weight gain. The most significant weight gain was observed in nails treated with 20% TA. In FIG. 5 shows the increase in nail weight in percent after a period of 20 hours at various concentrations of urea-H ₂ O ₂ . Percentages shown on the X axis indicate the total concentration of H ₂ O ₂ present (value ± standard deviation, n = 10). The concentration of urea-H ₂ O ₂ applied to the nails also affects weight gain and, thus, the permeability of the nail. The application of 20% urea-H ₂ O ₂ was significantly better than 10% in relation to the increase in the weight of the nail. However, nails treated with increasing concentrations of H ₂ O ₂ from 20% to 35% showed no significant difference (p> 0.05, ANOVA) in relation to weight gain.

In FIG. Figure 6 shows the increase in nail weight in percent after a period of 20 hours at various concentrations of DTT (value ± standard deviation, n = 10). Nails treated with 1% DTT showed a significant increase in weight (p> 0.05, ANOVA) compared to nails treated with 5% DTT, but further increases in concentration from 5 to 10% had no significant effect (p <0, 05, ANOVA). The greatest effect on the weight of the nail was observed at a concentration of DTT of 20%, which was almost twice the effect observed at a concentration of 10%.

Example 4: Effect of pH on penetration enhancers

Nail swelling studies:

In FIG. Figure 7 shows the increase in nail weight in percent after a 20-hour period with various TA salts (all 5% concentration). The numbers above the bars indicate the pH of each solution before application (value ± standard deviation, n = 10). Regardless of the type of thioglycolate salt used as a penetration enhancer, the increase in nail weight was significantly greater (p> 0.05, ANOVA) than in the control. Calcium thioglycolate caused the greatest increase in the weight of the nail and, therefore, facilitated the penetration of the greatest amount of fluid through the nail. The type of salt had a direct effect on the pH of the liquid applied to the nail. TA was used as a solution with a pH of 2.12, while calcium thioglycolate had a pH of 11.43. Thus, while all the compounds effectively enhanced fluid uptake by the nail, it was believed that pH> 7.1 was favorable in relation to the increase in the weight of the nail.

Examples 5-10 :

Materials and methods:

In Examples 5-10, the materials and methods used are described in detail below.

The results of measuring the redox potential:

An ion analysis redoximeter (Metrohm, UK) was used to determine the reduction potentials of 0.5 M aqueous compositions of compounds in deionized water against a stable reference electrode having a known potential (Ag / AgCl) at r.tp.

Terbinafine diffusion studies:

Samples of a human nail (total thickness) with a diameter of approximately 3 mm are placed between the two halves of the specially designed ChubTur ™ diffuser and clamped together. The sensor space is filled with a suitable buffer system, previously subjected to ultrasound in order to provide deposition conditions, and the cells are fixed on a Plexiglass stand mounted on a magnetic stirrer in a water bath maintained at 32 ° C. The contents of the sensor chamber are continuously mixed using small PTFE-coated magnetic push rods driven by an immersion magnetic stirrer. A known amount of the composition / solution of the drug is applied to the surface of the nail (an infinitely large dose), and at regular intervals, samples of the buffer are sampled from the sensor space, replaced with fresh medium for the sensor and examined for drug content using HPLC.

TurChub® Microbiological Tests:

Trichophyton rubrum was seeded onto a Saburo dextrose agar plate by carefully picking up the mycelium and spores using a sterile culture swab with a beveled culture medium and moving them to the surface of the agar. The cup was incubated at 25 ° C for 7 days. White spores on the surface of the cup are washed off with Ringer's solution (20 ml). The spore suspension is filtered through sterile gauze (Smith + Nephew, Propax, 7.5 cm × 7.5 cm, 8 layer gauze swab, BP Type 13) to remove the mycelium. Viable spores in suspension are counted and the spores are adjusted to approximately 1 × 10 ⁷ CFU / ml by diluting or concentrating the spores in a final volume of 20 ml, respectively.

Initially, distal nail pieces are obtained from volunteer toe nails, which have grown to a minimum length of 3 mm. All volunteers were required to not apply nail polish or gloss on their toenails for 6 months and not have any symptoms of the disease on their nails for 6 months. All volunteers were asked to remove the distal sections of the nail, using either scissors or standard nail clippers. No specific procedures are required, such as sterile removal or cleaning of nails. Then the nail pieces are placed in a suitable container, for example a plastic bag, a test tube, an envelope, etc., for example, placed in a small 8 ml vial per donor / dose, and labeled with any available parts. Samples are stored in the freezer until the right time.

Using scissors, the nail pieces are then cut into pieces that have a minimum size of 3 mm by 3 mm. The number of parts obtained from each nail depends entirely on the size of the initial sample, for example, only 1 or 2 parts can be obtained from the nail of the small toe, while from 3 to 5 parts can be obtained from the nail of the big toe. Pieces of the nail are immersed in 70% ethanol in an aqueous solution and mixed vortex for one minute. Then the ethanol solution is decanted and replaced with a fresh 70% ethanol solution and stirred vortex for another minute. Then, the ethanol solution is decanted and replaced with Ringer's solution, vortex-stirred for one minute and decanted and replaced with fresh Ringer's solution. This washing process with Ringer's solution is repeated a total of three times, replacing the washing solution in each phase. Upon completion of the washing process, the nail pieces are placed in a sterile Petri dish without a lid and dried in a chamber with a laminar flow of air for 30 minutes at room temperature. When the nail pieces are dried, they are placed in new small 8 ml bottles (separate bottle per donor, per serving). The thickness of the nails is also determined using a pair of calipers.

The distal pieces of the nail (the entire thickness of the nail) are then treated in a diffuser with a penetration enhancer system for 20 hours with thioglycolic acid, and then for 20 hours with urea H ₂ O ₂ .

Initially, TurChub® cells (lower and upper sections) are autoclaved at 121 ° C for 15 minutes. The nail sealant is also sterilized in 100% ethanol and air-dried in a laminar flow chamber before installing the nail / membrane sections. The nails are treated with a penetration enhancer, and then the lower half of the TurChub® cell is filled with the sealant with the dorsal side up. Then, a predetermined volume (calibrated for each cell) of the molten SDA agar (maintained at 56 ° C) is placed in the lower portion of the cell. After the agar accepts a fixed volume (50 μl) of the experiment, the T.rubrum organism in Ringer's solution is applied to the surface of the agar. Then contribute to the uniform distribution of the suspension of the organism on the surface of the agar by carefully swaying the cell from side to side. Excess fluid from the body suspension is removed from the cell using a syringe and a needle. After application to the cell of the body, a nail is set, and the upper funnel-shaped portion of the cell is added and fixed in position with the springs. The cells are designed so that, firstly, excess fluid (for example, from condensation) does not cause cross-contamination of the agar, but accumulates at the base of the cell; secondly, the cells are oriented in such a way as to avoid false positive results from loading to removing agar.

Approximately 100 μl of the active substance or composition is applied for 7 days next to the experimental organism T.rubrum. The effectiveness of the compositions is determined by measuring the zone of growth inhibition of T.rubrum in a TurChub ™ cell.

Example 5 : Measurement of the redox potential of penetration enhancers for nails

The redox potential of agents shown to enhance swelling of human nails is tested. It is believed that reducing agents will have -ve redox potentials, while oxidizing agents will have + ve redox potentials (when expressed, alternatively, as reducing potentials, the sign changes so that the reducing agents have + ve potential). The results in table 3 and in FIG. 8 (± standard deviation, n = 3) demonstrate that there are clear differences in the value of the redox potentials of different test agents.

Table 3 Values of redox potentials and pH of chemical agents (0.5 M concentration, n = 3) No. Chemical compound The average value of the redox potential (mV) Standard deviation PH average Standard deviation one Resorcinol 8.6 5.3 5.67 0.42 2 Cineol 67.2 100.5 5.91 0.30 3 Calcium thioglycolate -531.3 12.5 11.74 0.24 four p-coumaric acid 114.1 50.8 3,54 0.31 5 Thiouracil 53,4 18.9 6.28 0.08 6 Potassium persulfate 402.5 28.6 2.83 0.06 7 Glycolic acid 247.7 27.7 2.03 0.15 8 Oxalic acid 254.8 26.0 1.33 0.08 9 Hydroperite 349.7 5.3 2.72 0.31 10 Sodium thioglycolate -281.7 5.3 6.90 0.05 eleven Thioglycolic acid -132.8 13.8 2.13 0.02 12 Ammonium Thioglycolate -235.9 24.0 6.29 0.20 13 Hydrogen peroxide 277.3 48.3 6.21 0.31 fourteen Dtt -241.0 10.9 2.64 0.53 fifteen Cysteine -79.3 8.2 --- ---

Chemical agents that were highly reducing include salts of thioglycolic acid, with calcium thioglycolate, which has the lowest redox potential among all chemicals (-531.3 mV). The most powerful oxidizing agents were those with the highest redox potentials, which include hydroperite (UrCa-H ₂ O ₂ ), hydrogen peroxide (H ₂ O ₂ ), glycolic acid, oxalic acid and potassium persulfate.

From these redox potential data, it is clear that all combinations of agents that dramatically improved nail swelling (Example 2) were strong either oxidizing or reducing agents. In addition, the sequence of application of these agents is important, the greatest swelling of the nail in example 2 was observed when a reducing agent was initially applied to the nail.

Example 6 : Effect of the concentration of a compound enhancing penetration into the nail on the redox potential:

The effect of concentration on the redox potentials of the selected agents was investigated. These included thioglycolic acid, UTCa-H ₂ O ₂ , H ₂ O ₂ and DTT. The titration results are shown in tables 4-7.

Table 4 Average redox potentials and pH values TA at
concentrations from 0.001% to 20% w / w. (n = 3) TA concentration (% w / w) The average value of the redox potential Standard deviation PH average Standard deviation twenty% -128.7 0.1 1,61 0.04 10% -132.8 0.6 1.83 0.14 7.5% -132.7 0.1 1.76 0,03 5% -131.5 0.4 1.85 0.02 2.5% -126.5 0.5 1.71 0.55 one% -119.8 0.4 2.23 0.08 0.1% -97.4 1.4 2.66 0.13 0.05% -85.6 0.8 2,53 0.12 0.01% -67.5 1,2 3.05 0.12 0.005% -56.6 3.8 3.12 0.19 0.001% -47.7 5,4 2.97 0.32

Table 4 shows that at TA concentrations from 1 to 20%, there is no effect of concentration on the measured redox potential of the solution. At concentrations below these (0.001-1%), the redox potential steadily decreases with a decrease in concentration. A similar trend was observed when the redox potential of DTT was determined, and only at concentrations below 0.05% the redox potential of the solution decreased (table 5).

Table 5 Average redox potential and pH DTT at concentrations
from 0.005 to 12% w / w. (n = 2) DTT concentration (% w / w) The average value of the redox potential Standard deviation PH average Standard deviation 10 -193.1 11,45513 5.69 0.05 7.5 -209.65 16,33417 6.24 0.30 5 -201.25 16,05132 6.20 0.23 one -185.75 10.39447 7.01 0.06 0.1 -189.65 0.919239 7.59 0.09 0.05 -169.19 0.410122 7.80 0.12 0.01 -134.9 4,666905 8.00 0.02 0.005 -122.7 3,818377 8.07 0.45

A similar effect was noted with both H ₂ O ₂ and urea-H ₂ O ₂ . It turned out that at high concentrations from 5 to 17.5% of the last compound, no large differences in the redox potential were observed (table 6). However, with a decrease in the concentration of urea-H ₂ O ₂ below 5%, the redox potential and, thus, the oxidation energy decreased more rapidly. The pH of the solution also became more neutral as the concentration of the solution decreased. The critical concentration of H ₂ O ₂ at which the redox potential decreased was 20% (table 7).

Table 6 The average values of the redox potential and the pH of urea-H ₂ O ₂ at concentrations contained H ₂ O ₂ from 0.0025 to 17.5 H ₂ O ₂ wt./weight. (n = 3) The content of H ₂ O ₂ (% wt./weight.) The average value of the redox potential Standard deviation PH average Standard deviation 17.5 382.3 1.3 3.33 0.09 16.25 382.9 1,5 3.87 0.08 fifteen 380.3 0.8 4.04 0,03 10 377.9 0.5 4.23 0.12 5 372.3 1.4 5.10 0.11 0.5 293.6 11.2 5.31 0.33 0.05 208.1 5,0 6.81 0.16 0,025 199.5 3.4 6.56 0.04 0.005 191.3 5.2 6.17 1.00 0.0025 189.4 6.0 6.07 0.53

Table 7 Average redox potentials and pH values of H ₂ O ₂ at concentrations
from 0.005 to 35% w / w. (n = 3) The concentration of H ₂ O ₂ (% wt./weight.) The average value of the redox potential Standard deviation PH average Standard deviation 35 444.5 0.6 2,55 0.04 32,5 437.9 1.7 2.54 0.08 thirty 430.4 3.4 2.64 0.02 twenty 402.8 1.3 3.16 0.01 10 357.2 3,7 3.79 0.02 one 231.6 9.1 6.17 0,03 0.1 193.5 3.8 6.71 0.06 0.05 200,4 7.8 6.78 0.12 0.01 153.0 18,4 7.46 0.26 0.005 142.3 6.6 7.56 0.08

Example 7 : Effect of pH of a nail enhancement compound on redox potential:

To determine whether the pH of the solution has an effect on the redox potential of penetration enhancers, examine the pH range of two compounds: a 5% solution of TA and UrCa-H ₂ O ₂ with a H ₂ O ₂ content _of 15%. TAs are tested using a pH range of 2-11 (pH adjusted with NaOH solution). The urea-H ₂ O ₂ solution is tested at a pH of 2 to 10 (pH adjusted using HCl and NaOH solutions).

The redox potential of TA almost increased one order of magnitude over the tested pH range (table 8). In contrast, the redox potential of the UTCa-H ₂ O ₂ complex decreased at a constant rate until pH 10 was reached, after which a sharp drop in the redox potential was noted (table 9).

Table 8 The average redox potential of TA solutions at pH values ranging from 2 to 11 (n = 3) Estimated pH TA The average value of the redox potential Standard deviation 2 -70.4 20,4 3 -133.1 13.3 four -166.7 24.4 5 -179.5 33.7 6 -192.9 37.3 8 -272.3 49.4 10 -386.5 52.0 eleven -433.8 50.6

Table 9 The average redox potential of urea-H ₂ O ₂ solutions at
pH ranging from 2 to 10 (n = 3 ± standard deviation) The approximate pH value of urea-H ₂ O ₂ Redox potential (mV) Standard deviation 2 459.8 2.0 four 334.1 5.5 6 284.0 1.8 8 194.2 0.7 10 16.1 1,2

Example 8 : improved penetration of an example of a pharmaceutical compound

In FIG. Figure 9 graphically shows the diffusion of terbinafine through a human nail after pretreatment with either a 50:50 mixture of ethanol / water (control), urea-H ₂ O ₂ or thioglycolic acid (TA) separately, and these two penetration enhancers in combination. The results demonstrate the ability of a dual penetration enhancer system to increase the diffusion of a typical drug.

When the nails were kept in a 50:50 ethanol / water solution (control) after 20 hours and after application of the drug, only 44 ± 13 μg per cm ² penetrated through the nail after 214 hours. The application of an oxidizing agent, hydroperite, had no significant effect (p> 0.05, ANOVA) on the amount of drug penetrating the nail, allowing 47 ± 33 μg per cm ² to pass through the barrier. The application of thioglycolic acid to the nail prior to the application of terbinafine ensured the penetration of 861 ± 135 μg in cm ^{2 of the} drug, but this was less than with the sequential application of thioglycolic acid and urea-H ₂ O ₂ to terbinafine, since this ensured the penetration of 2308 ± 1332 μg per cm ² after 219 hours.

Example 9 : Effect of application time on the efficiency of a dual-penetration enhancer system:

In FIG. 10 graphically shows the diffusion of terbinafine through a human nail after pretreatment with thioglycolic acid (TA) and then urea-H ₂ O ₂ at various time intervals.

The use of a penetration enhancer system for either 30 minutes or 1 hour in both cases ensured penetration through the nail of less than 10 μg per cm ^{2 of the} drug. In contrast, when using a dual penetration enhancer system for 20 hours, each increased the amount of terbinafine penetrating the human nail to 3329 ± 1842 μg per cm ² after 315 hours.

Example 10 : Use of a penetration enhancer system to improve the effect of commercial antifungal compositions:

The effect of the dual penetration enhancer system on the penetration of amorolfine from Loceryl® commercial pharmaceutical varnish was tested using a microbiological test. The analysis evaluated the effectiveness of the composition, either individually or with pre-treatment or with thioglycolic acid, hydroperit, or both, against the predominant microorganism found in nails with onychomycosis, T. rubrum. Figure 11 shows a comparison of the average length of the inhibition zone (ZOI) when using the TurChub® cell system with T. rubrum, after applying various penetration enhancing systems to the pre-processed pieces of the distal nail of the entire thickness of the nail, and then applying 100 μl of the standard Loceryl varnish on the surface of each and incubation for seven days (n = 4 ± standard deviation).

In FIG. 11 shows that when amorolfin was either applied alone or after pretreatment of the nail for 20 hours with one reducing agent, thioglycolic acid, the growth of the microorganism was not prevented, that is, there was no detectable zone of inhibition. When an oxidizing agent, hydroperite, was used to pretreat the nail, a small zone of inhibition was detected. However, when both the oxidizing and reducing agent were sequentially applied to the nail before the antifungal drug, an inhibition zone of almost 2 cm was observed. These results emphasize that the application of both the oxidizing and reducing agents significantly enhances the microbiological destruction achieved by using a commercially available nail composition .

Claims (45)

1. A method of treating a pathological condition of nails in a patient in need of such treatment with a medicament, further using a first composition containing a reducing agent with a potential of more than +20 mV, and a second composition containing an oxidizing agent with a potential of less than -50 mV and separately separately sequentially on the nail of the specified patient, first the first composition is applied, then the second composition, the specified drug being included in the first composition, or in the second composition, or, optionally, in the composition of the third composition, which is a separate additional tool for compositions with reducing and oxidizing agents. 2. The method according to claim 1, in which a simple combination of these two compositions can lead to extreme reactions and in which each composition is individually pharmaceutically acceptable for application to the nails. 3. The method according to claim 1, wherein said reducing agent is selected from the group consisting of ammonium thioglycolate, calcium thioglycolate, sodium thioglycolate, thioglycolic acid and dithiothreitol, ascorbic acid, hydroquinone, mercaptoethanol, glutathione, L-cysteine, taurine, aminomethanesulfonic acid , cysteic acid, cysteinesulfinic acid, ethanedisulfonic acid, ethanesulfonic acid, homotaurin, hypothaurine, isethionic acid, mercaptoethanesulfonic acid, N-methyltaurine and their simple derivatives. 4. The method according to claim 3, in which the specified simple derivative is a salt. 5. The method according to claim 3, in which the specified reducing agent is thioglycolic acid or its derivative. 6. The method according to claim 1, wherein said oxidizing agent is selected from the group consisting of urea, hydrogen peroxide, potassium persulfate, thiouracil, p-coumaric acid, glycolic acid, oxalic acid, cineole, peroxidone, chlorine dioxide, ammonium dichromate, ammonium nitrate, ammonium perchlorate, ammonium permanganate, barium bromate, barium chlorate, barium peroxide, cadmium chlorate, calcium chlorate, calcium chromate, calcium perchlorate, chromium nitrate, cobalt nitrate, silver oxide, periodic acid and pyridine dichromate. 7. The method according to claim 1, wherein said oxidizing agent is hydrogen peroxide. 8. The method according to claim 1, wherein said oxidizing agent is an additional compound selected from hydrogen peroxide and urea. 9. The method according to claim 1, in which the drug is present in one or both of these compositions. 10. The method according to claim 1, in which the drug is present in the third composition. 11. The method according to claim 1, in which the compositions are liquid. 12. The method according to claim 1, in which the reducing agent is made in the form of an alkaline composition with a pH between 7 and 14. 13. The method according to item 12, in which the pH is between 8 and 13. 14. The method according to item 12, in which the pH is between 9 and 12. 15. The method according to claim 1, in which the pH of the oxidizing agent is between 1 and 7. 16. The method according to clause 15, in which the pH is between 2 and 5. 17. The method according to claim 1, in which the compositions are aqueous. 18. The method according to claim 1, comprising propylene glycol as a bulking agent. 19. The method according to claim 1, in which thioglycolic acid is conjugated to a drug. 20. The method according to claim 1, wherein said drug is selected from the group of antifungal drugs consisting of amorolfine, miconazole, ketoconazole, itraconazole, fluconazole, econazole, cyclopirox, oxyconazole, clotrimazole, terbinafine, naftifine, amphotericinazine, grizofericin, griz, flucytosine, nystatin and their pharmaceutically acceptable salts and esters. 21. The method according to claim 1, wherein said drug is terbinafine. 22. The method according to claim 1, wherein said drug is amorolfine. 23. The method according to claim 1, wherein the drug is selected from the group of antipsoriatic drugs consisting of corticosteroids, 5-fluorouracil, methotrexate, etretinate, cyclosporin, tacrolimus, and their derivatives. 24. The method according to claim 1, wherein said oxidizing agent has a reducing potential and wherein said reducing potential of 0.5 M of said oxidizing agent in deionized water, when determined against an Ag / AgCl reference electrode, is less than -50 mV. 25. The method according to paragraph 24, wherein said reduction potential is less than -100 mV. 26. The method according to paragraph 24, in which the specified recovery potential is less than -200 mV. 27. The method according to claim 1, in which the reducing agent has a reducing potential and wherein said reducing potential of 0.5 M of said reducing agent in deionized water, when determined against an Ag / AgCl reference electrode, is greater than +20 mV. 28. The method according to item 27, in which the specified recovery potential is greater than +50 mV. 29. The method according to item 27, in which the specified recovery potential is greater than +75 mV. 30. The method of claim 1, wherein said reducing agent is conjugated to said drug. 31. The method according to claim 1, wherein said oxidizing agent is conjugated to said drug. 32. The method according to claim 1, in which at least one of these compositions is selected from the group consisting of creams, ointments, gels, solutions, lotions, foams, mousses, sprays, pastes, dressings, powders, premixes, non-aqueous varnishes and waterborne varnishes. 33. The method according to claim 1, in which these compositions are separately selected from the group consisting of creams, ointments, gels, solutions, lotions, foams, mousses, sprays, pastes, dressings, powders, premixes, non-aqueous varnishes and aqueous varnishes. 34. The method according to claim 1, in which at least one of these compositions is a water-based gel. 35. The method according to claim 1, in which all of these compositions are water-based gels. 36. The method according to claim 1, in which at least one of these compositions is a water-based varnish. 37. The method according to claim 1, in which all of these compositions are water-based varnishes. 38. The method according to claim 1, wherein said first composition is applied at least 10 hours before said second composition. 39. The method according to claim 1, wherein said first composition is applied at least 15 hours before said second composition. 40. The method according to claim 1, wherein said first composition is applied at least 20 hours before said second composition. 41. The method according to claim 1, in which the specified third composition is applied within 1 hour after applying the specified composition of the oxidizing agent. 42. The method according to claim 1, in which the specified third composition is applied within 20 minutes after applying the specified second composition. 43. The method according to paragraph 41, wherein said third composition is applied immediately after application of the specified second composition. 44. A method of treating a nail infection in a patient in need of such treatment with a medicament, further using a first composition containing a reducing agent with a potential of more than +20 mV, and a second composition containing an oxidizing agent with a potential of less than -50 mV, moreover, separately sequentially on the nail of the specified patient is applied first the first composition, then the second composition, and the specified drug is included in the first composition, or in the second composition, or, neo optionally, in the composition of the third composition, which is a separate additional tool for compositions with reducing and oxidizing agents. 45. A kit comprising a first composition containing a reducing agent with a potential of more than +20 mV, a second composition containing an oxidizing agent with a potential of less than -50 mV, which are located separately, and a medicine for treating a pathological condition of the nails, wherein said medicine is included in the composition of the first composition, or the composition of the second composition, or, optionally, the composition of the third composition, which is a separate additional tool for compositions with a reducing and oxidizing agent and.

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